Contributory role of endothelium and voltage-gated potassium channels in apocynin-induced vasorelaxations

Objective: Although apocynin, the nicotinamide adenine dinucleotide phosphate oxidase inhibitor improves vascular function in hypertension, its specificity has been questioned. The present study examined whether apocynin-induced vasorelaxations involve endothelium and/or K channels in vascular cells. Methods: Aortas from Sprague-Dawley rats were suspended in organ baths for functional studies. Changes in intracellular calcium ([Ca ²⁺]_i) and nitric oxide ([NO]_i) in rat endothelial cells were detected by fluorescence imaging. Whole-cell voltage-gated K⁺(Kv) currents were recorded in vascular smooth muscle cells. Results: Apocynin-induced aortic relaxations were attenuated by the absence of endothelium, endothelial nitric oxide synthase inhibitor N ^G-nitro-L-arginine methyl ester, or nitric oxide-dependent soluble guanylate cyclase inhibitor 1H-[1,2,4] oxadiazolo [4,3-a] quinoxalin-1-one. 4-Aminopyridine (4-AP, voltage-gated potassium channels blocker) attenuated apocynin-induced relaxations, its combined treatment with 1H-[1,2,4] oxadiazolo [4,3-a] quinoxalin-1-one (ODQ) did not cause further inhibition. Apocynin increased [Ca²⁺]_i and [NO]_i in endothelial cells, which were abolished by 4-AP, indicating the involvement of voltage-gated potassium channels in endothelial cells. In addition, apocynin-stimulated increase in endothelial nitric oxide synthase phosphorylation at Ser ¹⁷⁷ depended on the presence of extracellular Ca²⁺. Notably, 4-AP also reduced apocynin-induced relaxations in the absence of endothelium and apocynin increased 4-AP-sensitive voltage-gated potassium channel currents in vascular smooth muscle cells. Conclusion: This study provides novel data showing that apocynin-induced relaxations of rat aortas are mediated by 4-AP-sensitive stimulation of [Ca²⁺]_i and [NO]_i increases in endothelium and by activation of voltage-gated potassium channels in vascular smooth muscle cells.